The invention concerns a method for the determination of pathogenic Listeria bacteria, a nucleic acid probe, as well as a protein which is suitable for the production of antibodies against pathogenic Listeria bacteria.
Listeria are a heterogeneous group of gram-positive bacteria and consist essentially of the species Listeria monocytogenes, L. innocua, L. welshimeri, L. seeligeri, L. ivanovii, L. grayi and L. murrayi. Only two of these species are pathogenic, namely L. monocytogenes for humans and animals and L. ivanovii for animals. Listeriosis usually manifests itself in humans as a bacterial meningitis and septicaemia as well as miscarriages and stillbirths in pregnant women. In sheep and cattle listeriosis manifests itself as miscarriage, encephalitis, septicaemia and mastitis. (N. Engl. J. Med. 308 (1983), 203-206, J. Infec. 15 (1987), 165-168, Linnan, M. J. et al., An investigation of listeriosis in Southern California 1985, in Courtieu, A. L. et al., (eds) Listeriose, Listeria, Listeriosis 1985-1986, University of Nantes).
Recently an increasing number of listeriosis diseases have been observed in humans, the cause of which is regarded to be due to the contamination of milk and cheese, above all soft cheese varieties, by Listeria bacteria. Therefore an examination of food for Listeria contamination is important and is obligatory for cheese in the USA.
A method for the determination of Listeria monocytogenes is known from Int. J. Food Microbiol. 4 (1987), 249-256 in which the microorganism is enriched selectively in a liquid medium at 4.degree. C., cultured on special agar plates and subsequently the biotype and serotype determined by a multitude of biochemical tests. This method is very laborious and time-consuming and it takes 14 days and longer before the end result is available. In addition the assessment is very difficult to carry out and is not suitable for routine investigation.
A method for the determination of Listeria bacteria is known from Appl. Environ. Microbiol. 53 (1987), 2256-2259 which is carried out by colony-hybridization with radioactively labelled DNA probes. Colony-hybridization in itself would be a suitable method for a rapid and simple analysis of the contamination of food by Listeria bacteria. However, a nucleic acid probe is necessary for this which hybridizes with all pathogenic Listeria bacteria (L. monocytogenes and L. ivanovii) but not with the other Listeria bacteria. DNA probes known up to now (listeriolysin 0, .beta.-listeriolysin; cf. Infection and Immunity 55 (1987), 3225-3227 and Applied Environmental, Microbiology 53 (1987), 2256-2259) are too non-specific and therefore result in an unacceptable number of false positive and false negative results.
The object of the present invention is therefore to provide a nucleic acid with which specific hybridization tests for pathogenic Listeria bacteria are possible as well as proteins derived from this which are suitable as immunogens for the production of antibodies against pathogenic Listeria bacteria.